The goal of this proposal is to obtain new insights into the role of proteinases and their inhibitors during cerebral ischemia, and to relate these findings to the potential development of neuroprotective strategies. The Holland Laboratory of the American Red Cross is the primary research facility of the American Red Cross and is ideally suited to pursuit these goals. The Principal Investigator is a Stroke Neurologist trained at Georgetown University Hospital who wants to develop a career as a physician-scientist in the field of cerebrovascular disease. This requires the development of skills in basic science research that will allow future work to range "from the bench to the bedside." The Mentored Clinician Scientist award will permit the applicant to develop a career as an independent basic researcher under the supervision of a mentor that has been at the Holland Laboratory since 1995, and has extensive experience in the molecular aspects of proteinases-inhibitor interaction both in vitro and in vivo. The principal focus of this study is the neuroprotective role of the serine proteinase inhibitor (serpin) neuroserpin during cerebral ischemia. Neuroserpin is a recently identified member of the serpin gene family, that is primarily localized to the brain, where it appears to act as a specific inhibitor of tissue type plasminogen activator (tPA). During acute occlusive stroke, when the blood supply to the brain is interrupted, there is a densely ischemic area of necrotic tissue, surrounded by a potentially recoverable zone, known as "ischemic penumbra." Preliminary studies indicate that endogenous neuroserpin expression is increased in the ischemic penumbra, and that administration of neuroserpin after stroke decreases infarct volume. The reduction in infarct size correlates with a decrease in neuronal apoptosis within the ischemic penumbra, and with an apparent reduction in degradation of the vascular basement membrane. The studies described herein will focus on the following issues: What is the effect of neuroserpin on the evolution of the ischemic penumbra? Is neuroserpin a naturally occurring neuroprotective agent? What is the mechanism by which neuroserpin treatment decreases infarct volume and neuronal apoptosis? Does neuroserpin preserve the integrity of the basement membrane and thus the blood-brain barrier after stroke? Is neuroserpin's only target tPA or does administration of neuroserpin affect other proteinases such as urokinase plasminogen activator (uPA) or matrix metalloproteinases (MMPs)? Together, these studies should help to delineate many important functions of neuroserpin and provide a better understanding of its role during cerebral ischemia.